Project Summary Brain-wide correlation of single-cell firing properties to patterns of gene expression Every neuron expresses thousands of proteins which, through their interactions with each other and with other small molecules, imbue the neuron with its functional properties. The primary function of each neuron is to receive synaptic inputs, to produce electrical spikes, and to release neurotransmitters to its downstream neighbors. Neurons show widely varying patterns of gene expression, and also show widely varying patterns of electrical spiking. However, the mapping of gene expression onto spiking behavior is unknown. Due to the complexity of the interactions within a cell, it is not currently possible to predict the electrical spiking properties of a cell from its pattern of gene expression. The present proposal seeks to develop tools to record gene expression and spiking patterns in thousands of neurons throughout a brain slice. These measurements will reveal the correlations between genes and neural function, which in turn will suggest hypotheses about how modifications to specific genes might affect neural function. The proposal seeks to combine two new technologies: all-optical electrophysiology (`Optopatch') and multiplexed error-robust fluorescence in situ hybridization (`MERFISH'). A novel instrument will be developed, capable of recording Optopatch data on thousands of neurons across a brain slice. A set of protocols will be developed to perform sequential Optopatch and MERFISH profiling on the same brain slice, and to correlate the two datasets with single-cell resolution. These techniques will then be applied to map firing properties and gene expression across rodent brain slices and in zebrafish.